Polycystic Ovary Syndrome (PCOS) is an endocrinopathy of unknown etiology that affects as many as 8% of the female population and is the most common cause of female infertility. The chronic anovulation and hyperandrogenism characteristic of PCOS is promulgated by insulin resistance and the resultant hyperinsulinemia in the majority of affected individuals. The insulin resistance predisposes those with the disorder to type 2 diabetes and is a known risk factor for early onset of cardiovascular disease. Tumor necrosis factor alpha (TNF-alpha), an inflammatory cytokine produced by adipose tissue and activated mononuclear cells (MNC) has been implicated as a mediator of insulin resistance in PCOS. The proposed research will involve the perspective study of 40 reproductive age women (18-40 years). Twenty of the women will have PCOS (10 normal weight and 10 obese) and 20 will be weight-matched ovulatory controls. The central hypothesis is that a physiologic carbohydrate challenge triggers excessive TNF-alpha release from MNC of women with PCOS. The specific aims are: 1) to examine TNF-alpha release from MNC of women with PCOS in response to glucose stimulation; 2) to examine the inflammation pathway in MNC of women with PCOS in response to glucose stimulation. The approach involves the quantification of TNF-alpha release from MNC culture supernatants of women with PCOS before and after in vivo glucose exposure during a standard oral glucose challenge test and under euglycemic and hyperglycemic conditions in vitro using cultured MNC isolated from fasting blood samples. The inflammatory response of MNC in women with PCOS will also be evaluated before and after in vivo glucose challenge by measuring [reactive oxygen species] ROS generation, expression of protein markers of the inflammation pathway and activated nuclear factor kappa B (NFkB). It is our expectation that the approach used in this study will demonstrate excessive MNC-derived TNF-alpha release and a pronounced inflammatory response following glucose stimulation. These results will be significant in that they will identify the MNC as an additional source of excess TNF-alpha in PCOS aside from adipose tissue, and the inflammatory mechanism responsible for excessive TNF-alpha release from MNC. This will lead to important advances in the therapy of PCOS designed to ameliorate insulin resistance by reducing inflammation. It will also provide precedence for future studies to determine the molecular mechanism responsible for the sensitivity of MNC to postprandial hyperglycemia in women with PCOS that places these individuals in a proinflammatory state. [unreadable] [unreadable] [unreadable]